RNA-Seq of HepG2 cell line treated with AT7519 200nM for 24h
mRNA was purified from total RNA using poly-T magnetic beads. Fragmentation was carried out using divalent cations under elevated temperature in First Strand Synthesis Reaction Buffer. First strand cDNA was synthesized using random hexamer primer and M-MuLV Reverse Transcriptase. Second strand cDNA synthesis was performed using DNA Polymerase I and RNase H. Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities. After adenylation of 3 ends of DNA fragments, adaptors with hairpin loop structure were ligated. Library fragments were purified with AMPure XP. Then PCR was performed with Phusion High-Fidelity DNA polymerase, Universal PCR primers and Index (X) Primer. PCR products were purified.